Related Experiment Videos
Screening of combinatorial peptide libraries: identification of ligands for affinity purification of proteins using a
K Mondorf1, D B Kaufman, R G Carbonell
1Department of Chemical Engineering, North Carolina State University, Raleigh 27695-7905, USA.
Abstract:
Peptides deduced from peptide libraries may serve as affinity ligands for protein purification. Identification of a ligand that binds the protein of interest depends highly on the screening method used. One approach which offers simple and direct detection involves screening a solid-phase peptide library against a radiolabeled target protein. We have developed a radiological screening method, using 14C as a radioactive label, that offers high resolution and sensitivity. Less than 100 DPM/bead are detectable after a one-day exposure using autoradiography. The validity of the technique was illustrated by screening a solid-phase hexameric-peptide library spiked with YNFEVL-beads against 14C-labeled ribonuclease S-protein. For this particular system, the amount of protein bound to a single bead was estimated to be in the femtomolar range with a peptide:protein ratio of 500:1. Finally, a portion of the library was screened against 14C-labeled fibrinogen. Three peptides deduced from the library, WQEHYN, WQETYQ, and YENYGY, purified fibrinogen from a mixture with albumin.